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Sparked by the isolation of graphene in 2004, the research community has developed a family of 2D materials with distinct functionalities, enabling rapid demonstrations of entirely 2D devices with applications in energy, electronics, sensors and medicine. The desire to capitalize on the fantastic properties of 2D materials motivates ongoing efforts to synthesize high-quality 2D materials on large-scales. Epitaxial deposition of 2D materials on single-crystal substrates yields large-scale production of 2D materials without compromising crystal quality. However, 2D material properties are altered depending on the strength of the interactions with the substrate. The strength of these interactions, from weak van der Waals attractions to strong covalent bonds, are further revealed by local perturbations to the vertical atomic structure of the interface. For my dissertation research, I have employed high-resolution synchrotron X-ray characterization techniques to probe the interface between 2D materials and their growth substrate with atomic precision on wafer-scales. My dissertation primarily focuses on the interface of elemental 2D materials and single-crystal substrates, specifically 2D boron (borophene) on Ag(111) and graphene on Ge(110). The primary objective of these measurements is to resolve outstanding questions about the strength of interactions and resulting structure of these interfaces. Specifically, employing X-ray standing wave-enhanced photoelectron spectroscopy, I reveal borophene to be a highly planar atomic layer of B atoms by constructing a chemically-resolved vertical structure of borophene with sub-Å precision. Next, I show that epitaxial graphene (EG) atop Ge(110), upon annealing in ultra-high vacuum (UHV), induces a novel reconstruction of the underlying Ge(110) surface. I preform a detailed documentation of this structure using a combination of high-resolution X-ray reflectivity and grazing-incident X-ray diffraction. Finally, I probe the registry of van der Waals epitaxial materials using a combination of grazing-incident X-ray scattering measurements. Overall, the work presented herein reveals how distinct X-ray surface science techniques can be employed to resolve outstanding questions about new 2D material interfaces